A2 Β-casein Research Articles

Comparison of the impact of bovine milk β-casein variants on digestive comfort in females self-reporting dairy intolerance: a randomized controlled trial

Lactose malabsorption (LM) is a major cause of digestive discomfort from dairy products. Recently, a role for bovine β-casein A1 has been proposed. We examined whether there are distinct symptoms of digestive discomfort due to either lactose or differing bovine β-casein types. Women (n=40; age: 25.2 ± 0.5 y) with self-reported varying dairy tolerance underwent a 50-g lactose challenge. Based on postchallenge LM and digestive discomfort, participants were classified as either lactose intolerant (LI; n=10, self-reported intolerant, diagnosed lactose intolerant), nonlactose dairy intolerant (NLDI; n=20, self-reported intolerant, diagnosed lactose tolerant), or dairy tolerant (DT; n=10, self-reported tolerant, diagnosed lactose tolerant). In a double-blinded randomized sequence, participants consumed 750 mL conventional milk (CON; containing A1 and A2 β-casein and lactose), a2 Milk (A2M; exclusively containing A2 β-casein with lactose), or lactose-free conventional milk (LF-CON; containing A1 and A2 β-casein without lactose). Subjective digestive symptoms and breath hydrogen (measuring LM) were recorded regularly over 3 h, and further ad hoc digestive symptoms over 12h. LI subjects experienced prolonged digestive discomfort with CON milk. A2M reduced (P<0.05) some symptoms (nausea: A2M 8 ± 3 mm compared with CON 15 ± 3mm; fecal urgency: A2M 4 ± 1 compared with CON 10 ± 3 mm), and attenuated the rise in breath hydrogen over 3 h, relative to CON milk (A2M 59 ± 23 compared with CON 98 ± 25 ppm at 150 min; P<0.01). In contrast, NLDI subjects experienced rapid-onset, transient symptoms (abdominal distension, bloating, and flatulence) without increased breath hydrogen, irrespective of milk type. In LI individuals, LM and digestive comfort with lactose-containing milks was improved with milk containing exclusively A2 β-casein. Furthermore, self-reported dairy intolerance without LM (NLDI) is characterized by early-onset digestive discomfort following milk ingestion, irrespective of lactose content or β-casein type. This trial was registered at www.anzctr.org.au as ACTRN12616001694404.

Comparative Evaluation of Antioxidant Potential of In-Vitro Digested Cow Milk Derived A1 and A2 β Casein Variants Using Different Proteases

This study was designed to investigate the antioxidant activity of A1 and A2 β-casein variant with time by applying different enzyme. Pepsin, Trypsin, Alcalase and combination of Pepsin-Trypsin were used for hydrolysis of A1 and A2 β-casein for the duration of 1, 2, 3 and 24 h. All antioxidant parameter including DPPH, ABTS radical-scavenging activity and reducing power assay increasing gradually with time. Enzyme Pepsin-Trypsin combination followed by Alcalsae display comparatively higher antioxidant activity. Among β-casein variant A2 showed relatively higher antioxidant potential over all the entire duration of time but the difference among the A1 and A2 variants was not significant to arrive at a substantial scientific conclusion. It can be concluded from the study that antioxidant potential of the milk depends upon factors such as duration of hydrolysis and enzyme used, during hydrolysis and not alone on the fact that whether the milk is A1 or A2 in nature.

Manufacture and Physicochemical Properties of Chitosan Oligosaccharide/A2 β-Casein Nano-Delivery System Entrapped with Resveratrol.

The purposes of this research were to form chitosan oligosaccharide (CSO)/A2 β-casein nano-delivery systems (NDSs) and to investigate the effects of production variables, such as CSO concentration levels (0.1%, 0.2%, and 0.3%, w/v) and manufacturing temperature (5°C, 20°C, and 35°C), on the production and physicochemical characteristics of CSO/A2 β-casein NDSs to carry resveratrol. The morphological characteristics of CSO/A2 β-casein NDSs were assessed by the use of transmission electron microscopy (TEM) and particle size analyzer. High-performance liquid chromatography (HPLC) was applied to determine the entrapment efficiency (EE) of resveratrol. In the TEM images, globular-shaped particles with a diameter from 126 to 266 nm were examined implying that NDSs was successfully formed. As CSO concentration level was increased, the size and zeta-potential values of NDSs were significantly (p<0.05) increased. An increase in manufacturing temperature from 5°C to 35°C resulted in a significant (p<0.05) increase in the size and polydispersity index of NDSs. Over 85% of resveratrol was favorably entrapped in CSO/A2 β-casein NDSs. The entrapment efficiency (EE) of resveratrol was significantly (p<0.05) enhanced with an increase in manufacturing temperature while CSO concentration level did not significantly affect EE of resveratrol. There were no significant (p<0.05) changes observed in the size and polydispersity index of NDSs during heat treatments and storage in model milk and yogurt indicating that CSO/A2 β-casein NDSs exhibited excellent physical stability. In conclusion, the CSO concentration level and manufacturing temperature were the crucial determinants affecting the physicochemical characteristics of CSO/A2 β-casein NDSs containing resveratrol.

An idea to explore: Genotyping bull sperm to introduce basic molecular biology techniques in an animal science course.

The laboratory exercise described here aims to provide a relevant context for learning basic DNA techniques in an introductory animal science course at tertiary level. In two 4-hr laboratory sessions, students assess the suitability of bulls for inclusion in a gene-assisted selection program for A2 β-casein by genotyping commercial bull sperm. Sperm cells are lysed to extract the genomic DNA, and PCR with primers for the β-casein gene is performed. Using the principle of amplification-created restriction sites, restriction digestion with TaqI can be used to distinguish between the A1 allele and the A2 allele of the gene. Cut PCR amplicons are separated by gel electrophoresis to evaluate the genotype of each bull. Students then write a diagnostic report with accompanying letter to their fictional client, explaining the DNA test, and interpreting the results. © 2019 International Union of Biochemistry and Molecular Biology, 47(6):708-711, 2019.

Effects of Conventional Milk Versus Milk Containing Only A2 β-Casein on Digestion in Chinese Children: A Randomized Study.

ABSTRACTObjectives:In this study, we hypothesized that replacing conventional milk, which contains A1 and A2 β-casein proteins, with milk that contains only A2 β-casein in the diet of dairy or milk-intolerant preschoolers (age 5 to 6 years) would result in reduced gastrointestinal symptoms associated with milk intolerance, and that this would correspond with cognitive improvements.Methods:This randomized, double-blind, crossover study aimed to compare the effects of 5 days’ consumption of conventional milk versus milk containing only A2 β-casein on gastrointestinal symptoms, as assessed via visual analog scales, average stool frequency and consistency, and serum inflammatory and immune biomarkers in healthy preschoolers with mild-to-moderate milk intolerance. The study also aimed to compare changes in the cognitive behavior of preschoolers, based on Subtle Cognitive Impairment Test scores.Results:Subjects who consumed milk containing only A2 β-casein had significantly less severe gastrointestinal symptoms as measured by visual analog scales, reduced stool frequency, and improvements in stool consistency, compared with subjects consuming conventional milk. There were significant increases from baseline in serum interleukin-4, immunoglobulins G, E, and G1, and beta-casomorphin-7 coupled to lower glutathione levels, in subjects consuming conventional milk compared with milk containing only A2 β-casein. Subtle Cognitive Impairment Test analysis showed significant improvements in test accuracy after consumption of milk containing only A2 β-casein. There were no severe adverse events related to consumption of either milk product.Conclusions:Replacing conventional milk with milk containing only A2 β-casein reduced gastrointestinal symptoms associated with milk intolerance in Chinese preschool children, with corresponding improvements in aspects of cognitive performance.

Genetic association analysis reveals significant effect of β-casein A1/A2 loci on production & reproduction traits in Frieswal crossbred cows

ABSTRACT Milk β-casein has attracted extensive research interest in the dairy industry and animal breeding as a source of high-quality protein for human diet; however, adverse health effects have been ascribed to A1 β-casein variant consumption. A study was conducted to genotype the polymorphism in milk β-casein gene and examines the potential association between β-casein genotypes and milk production and reproduction traits in population of Frieswal cows. β-Casein A1/A2 genotype had significant effect (P ≤ 0.05) on several production traits including Total Milk Yield (TMY) (P ≤ 0.05), Milk Milk yield 300 days (MY300) (P ≤ 0.05), Lactation Length (LL) (P ≤ 0.01) and Peak Yield (PY) (P ≤ 0.01). β-Casein A1A1 genotype cows had significantly lower TMY, MY300, PY and DP as compared to A1A2 and A2A2 genotypes. A1A2 genotype had highest TMY, MY300, PY and DP; however, A2A2 and A1A2 genotypes were not significantly different from each other. The results pointed towards the superiority of A2 β-casein variant for several production traits in crossbreds. Further, with higher price of A2 milk, the inclusion of β-casein A1/A2 genotype in the selection of crossbreds offers higher profitability from dairy enterprise as well as provides a nutritional wellness product for consumers.

Milk A1 β-casein and health-related outcomes in humans: a systematic review.

Various epidemiological studies suggest a positive association between exposure to cow's milk A1 β-casein protein and risk for noncommunicable chronic diseases. The consumption of A2 cow's milk is increasing, likely because A2 milk is postulated to have positive effects on digestive health. A systematic review was conducted to investigate associations between A1 β-casein and health-related outcomes in humans. Five electronic databases, 3 clinical trial registries, and the internet were searched systematically. Using predefined inclusion criteria, 2 authors independently selected studies investigating the effect of A1 β-casein or β-casomorphin-7 intake/exposure on any health-related outcome in humans. Discrepancies were resolved by consensus. Two authors independently extracted data and assessed risk of bias. The certainty of evidence per outcome was evaluated using the GRADE approach. Discrepancies were resolved by consensus. Fifteen randomized controlled trials, 2 case-control studies, and 8 ecological studies were included. Most randomized controlled studies and case-control studies investigating a potential effect on various outcomes were based on intermediate markers and found no significant difference between the 2 milk types. In contrast, most ecological studies reported that population-level A1 β-casein exposure is associated with adverse health outcomes. The certainty of the evidence for the included outcomes, as assessed by the GRADE approach, was rated as moderate for digestive symptoms and as low to very low for all other outcomes. Human-based evidence from clinical trials and epidemiological studies published prior to October 2017 provides moderate certainty for adverse digestive health effects of A1 β-casein compared with A2 β-casein but low or very low certainty for other health effects. These conclusions may change in the future, given the emergent nature of this topic and the ongoing research in this area. PROSPERO registration number CRD42016043795.

DESARROLLO DE UNA FÓRMULA INFANTIL ALTA EN ALFA-LACTALBUMINA CON SOLAMENTE BETA-CASEINA A2 UTILIZANDO SECADO POR ASPERSIÓN, DISEÑADA PARA ASEMEJAR LA COMPOSICIÓN DE LA PROTEÍNA DE LA LECHE HUMANA

In this work, an infant formula enriched in α-lactalbumin and whose β-casein fraction contains only A2 β-casein was development by spray drying. Experimental infant formula (EF) was designed to closely resemble the protein composition of human milk. Response surface method (RSM) with two factors: total solids and inlet air temperature was used for the optimization of the spray drying process. Response variables were solubility, moisture content, luminance and peroxide value. To define optimal drying conditions, a stability test was performed for two treatments (120 °C, 40 wt. % and 170 °C, 30 wt. %), which were obtained from the RSM. The treatment with 170 °C and 30 wt. % concentration of total solids was selected as the best alternative for the spray drying process. The protein composition in the EF was 37.1% casein, 23.1% α-lactalbumin and 21.3% β-lactoglobulin. A decrease of 6 % in β-lactoglobulin increased the relative proportion of α-lactalbumin, which favoured the similarity of the EF to breast milk. The homology of the β-caseins with the A2 β-casein isoform was 99.82%. &nbsp

Sequence Characterisation and Genotyping of Allelic Variants of Beta Casein Gene Establishes Native Cattle of Ladakh to be a Natural Resource for A2 Milk

Bovine milk is regarded as nature's perfect food due to presence of vital nutrients. However some peptides are generated after proteolytic digestion of β-casein that have opioid properties and may increase the risk of chronic diseases. There are 13 genetic variants of bovine beta-casein; out of these A1 and A2 are the most common in dairy cattle breeds. The A1 and A2 variants differ only at position 67, which is histidine in A1 or proline in A2 milk. Earlier published reports have indicated that A1 β casein could be responsible for several health disorders like diabetes, coronary heart disease etc. while A2 β-casein is generally considered safe for human consumption. In the present study, an effort was made to sequence characterize β casein gene and identify allelic distribution of A1A2 alleles in native cattle of Ladakh region adapted to high altitude and low oxygen condition. The data showed 2 non-synonymous variations in coding region, while 5’UTR was completely conserved. The 3’UTR showed 2 more variations in Ladakhi samples. Further, the genotyping in 85 Ladakhi cattle for A1A2 alleles revealed that in Ladakhi cattle, A2 allele is predominantly present as reported for some of the other Indian breeds. The frequency of A2 allele was 0.90 and frequency of A2A2 genotype was found to be 0.79 in Ladakhi cattle. The present data strongly indicate that local cattle of Ladakh with higher frequency of A2 allele and A2A2 genotype is natural resource for A2 milk. Systematic efforts should be made for long term conservation and genetic improvement of this invaluable genetic resource of Ladakh.

Hypoallergenic and Physicochemical Properties of the A2 β-Casein Fractionof Goat Milk.

Goat milk has a protein composition similar to that of breast milk and contains abundant nutrients, but its use in functional foods is rather limited in comparison to milk from other sources. The aim of this study was to prepare a goat A2 β-casein fraction with improved digestibility and hypoallergenic properties. We investigated the optimal conditions for the separation of A2 β-casein fraction from goat milk by pH adjustment to pH 4.4 and treating the casein suspension with calcium chloride (0.05 M for 1 h at 25°C). Selective reduction of β-lactoglobulin and αs-casein was confirmed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis and reverse-phase high-performance liquid chromatography. The hypoallergenic property of A2 β-casein fraction was examined by measuring the release of histamine and tumor necrosis factor alpha from HMC-1 human mast cells exposed to different proteins, including A2 β-casein fraction. There was no significant difference in levels of both indicators between A2 β-casein treatment and the control (no protein treatment). The A2 β-casein fraction is abundant in essential amino acids, especially, branched-chain amino acids (leucine, valine, and isoleucine). The physicochemical properties of A2 β-casein fraction, including protein solubility and viscosity, are similar to those of bovine whole casein which is widely used as a protein source in various foods. Therefore, the goat A2 β-casein fraction may be useful as a food material with good digestibility and hypoallergenic properties for infants, the elderly, and people with metabolic disorders.

Clinical evaluation of glutathione concentrations after consumption of milk containing different subtypes of β-casein: results from a randomized, cross-over clinical trial.

This study reports the plasma glutathione concentrations in a double-blind, randomized, controlled, 2 × 2 cross-over study in which healthy participants consumed conventional milk (2 × 250 mL per day) containing both A1 and A2 types of β-casein, or milk containing only A2 type β-casein. Beta-casomorphin-7 (BCM-7), a peptide uniquely derived from the A1 type of β-casein, was previously reported to downregulate glutathione expression in human gut epithelial and neuronal cell lines by limiting cysteine uptake. The current human study demonstrates that consumption of milk containing only A2 β-casein was associated with a greater increase in plasma glutathione concentrations compared with the consumption of milk containing both β-casein types, and did not increase plasma BCM-7 concentrations compared with the washout diet in the study participants. Thus, milk containing only A2 β-casein and not A1 β-casein has the potential to promote the production of the antioxidant glutathione in humans.Clinical Trial RegistrationClinicalTrials.gov; identifier: NCT02406469 Electronic supplementary materialThe online version of this article (doi:10.1186/s12937-016-0201-x) contains supplementary material, which is available to authorized users.

Structural differences between bovine A1 and A2 β-casein alter micelle self-assembly and influence molecular chaperone activity

Within each milk protein there are many individual protein variants and marked alterations to milk functionality can occur depending on the genetic variants of each protein present. Bovine A1 and A2 β-casein (β-CN) are 2 variants that contribute to differences in the gelation performance of milk. The A1 and A2 β-CN variants differ by a single AA, the substitution of histidine for proline at position 67. β-Casein not only participates in formation of the casein micelle but also forms an oligomeric micelle itself and functions as a molecular chaperone to prevent the aggregation of a wide range of proteins, including the other caseins. Micelle assembly of A1 and A2 β-CN was investigated using dynamic light scattering and small-angle X-ray scattering, whereas protein functionality was assessed using fluorescence techniques and molecular chaperone assays. The A2 β-CN variant formed smaller micelles than A1 β-CN, with the monomer–micelle equilibrium of A2 β-CN being shifted toward the monomer. This shift most likely arose from structural differences between the 2 β-CN variants associated with the adoption of greater polyproline-II helix in A2 β-CN and most likely led to enhanced chaperone activity of A2 β-CN compared with A1 β-CN. The difference in micelle assembly, and hence chaperone activity, may provide explain differences in the functionality of homozygous A1 and A2 milk. The results of this study highlight that substitution of even a single AA can significantly alter the properties of an intrinsically unstructured protein such as β-CN and, in this case, may have an effect on the functionality of milk.

Dietary A1 β-casein affects gastrointestinal transit time, dipeptidyl peptidase-4 activity, and inflammatory status relative to A2 β-casein in Wistar rats

We compared the gastrointestinal effects of milk-based diets in which the β-casein component was either the A1 or A2 type in male Wistar rats fed the experimental diets for 36 or 84 h. Gastrointestinal transit time was significantly greater in the A1 group, as measured by titanium dioxide recovery in the last 24 h of feeding. Co-administration of naloxone decreased gastrointestinal transit time in the A1 diet group but not in the A2 diet group. Colonic myeloperoxidase and jejunal dipeptidyl peptidase (DPP)-4 activities were greater in the A1 group than in the A2 group. Naloxone attenuated the increase in myeloperoxidase activity but not that in DPP-4 activity in the A1 group. Naloxone did not affect myeloperoxidase activity or DPP-4 activity in the A2 group. These results confirm that A1 β-casein consumption has direct effects on gastrointestinal function via opioid-dependent (gastrointestinal transit and myeloperoxidase activity) and opioid-independent (DPP-4 activity) pathways.

Lower Consumption of Cow Milk Protein A1 β-Casein at 2 Years of Age, Rather than Consumption among 11- to 14-Year-Old Adolescents, May Explain the Lower Incidence of Type 1 Diabetes in Iceland than in Scandinavia

Aim: To compare the consumption of the cow milk proteins A1 and B β-casein among children and adolescents in Iceland and Scandinavia (Norway, Denmark, Sweden and Finland) as this might explain the lower incidence of type 1 diabetes (per 100,000/year, 0–14 years) in Iceland. Methods: The consumption of A1 β-casein in each country among 2- and 11- to 14-year-old children was calculated from results on food intake and on cow milk protein concentration. The consumption values were then compared and evaluated against the incidence of type 1 diabetes. Results: There was a significant difference between the consumption of A1 (p = 0.034) as well as the sum of A1 and B (p = 0.021) β-casein in Iceland and Scandinavia for 2-year-old children. In the same age group, consumption of A1 β-casein correlated with the incidence of type 1 diabetes in the countries (r = 0.9; p = 0.037). No significant difference in consumption of A1 or the sum of A1 and B β-casein was found for 11- to 14-year-old adolescents. Conclusion: This study supports that lower consumption of A1 β-casein might be related to the lower incidence of type 1 diabetes in Iceland than in Scandinavia. Additionally it indicates that consumption in young childhood might be of more importance for the development of the disease incidence than consumption in adolescence.

A comparison of the effects of A1 and A2 β-casein protein variants on blood cholesterol concentrations in New Zealand adults

β-Casein is a cow's milk protein that occurs predominantly in two forms, A1 and A2. Epidemiological evidence suggests that per capita consumption of β-casein A1 is associated with national mortality rates from ischaemic heart disease. A biological mechanism was proposed after rabbits fed diets containing β-casein A2 had lower serum cholesterol concentrations and less aortic intimal thickening than rabbits fed β-casein A1. We tested whether β-casein A1 and A2 variants differentially affect plasma cholesterol concentrations in humans. In a randomised crossover trial of two four-and-a-half week periods without washout, 62 participants replaced all dairy products in their diet with 500 mL of low-fat milk and 28 g of full-fat cheese that differed in the proportion of β-casein A1 and A2 variants. Duplicate blood samples were taken on non-consecutive days at the end of each treatment period from 55 people who completed the study. Mean (S.D.) plasma total, low-density and high-density lipoprotein cholesterol concentrations were 5.60 (0.77), 3.73 (0.70) and 1.26 (0.34) mmol/L after the A1 diet and 5.63 (0.81), 3.75 (0.75) and 1.27 (0.37) mmol/L after the A2 diets. We found no evidence that dairy products containing β-casein A1 or A2 exerted differential effects ( P > 0.05) on plasma cholesterol concentrations in humans.

A multi-centre, blinded international trial of the effect of A(1) and A(2) beta-casein variants on diabetes incidence in two rodent models of spontaneous Type I diabetes.

The diabetes-inducing potential of cows' milk is still debated and there is no consensus on the diabetogenicity of individual milk proteins. A(1)-beta-casein has been associated with increased diabetes frequency in ecological studies and in NOD mice. Our aim was to ascertain whether A(1)-beta-casein was more diabetogenic than A(2) and to test the diabetogenicity of a milk-free diet in animals representing different forms of spontaneous Type I (insulin-dependent) diabetes mellitus. Defined diets were coded and shipped to laboratories in New Zealand (NOD/NZ), Canada (BB) and the UK (NOD/Ba). Base diets were Pregestimil (PG) and ProSobee (PS). Purified fractions of whole casein (WC), A(1)or A(2)-beta-casein were added at 10%. A milk-free, wheat-predominant, NTP-2000 diet was the control. Animals were fed from weaning up to 150 or 250 days, and insulitis, diabetes frequency and expression of pancreatic cytokines were assessed. Diabetes incidence was highest in three locations in animals fed NTP-2000. PG and PS diets were protective except for NOD/Ba mice fed PG+WC where incidence was similar to NTP-2000. A(1) and A(2) diets were protective in both models, but A(1) beta-casein was slightly more diabetogenic in PS-fed BB rats. The New Zealand study was confounded by an infection. A milk-free, wheat-predominant diet was highly diabetogenic in three widely separate locations in both animal models. A previous result that A(1) beta-casein was more diabetogenic than A(2) beta-casein in NOD mice was not confirmed; both beta-casein variants were protective in BB rats and NOD mice. Whole Casein promoted diabetes in NOD/Ba but protected BB showing that unique diabetes haplotypes react differently to dietary proteins. A(1)- was more diabetogenic than A(2)-beta-casein only in PS-fed BB rats. Neither the analysis of insulitis nor of pancreatic cytokine gene expression showed a difference between A(1) or A(2) beta-casein fed animals. Milk caseins are unlikely to be exclusive promoters of Type I diabetes, but could enhance the outcome of diabetes in some cases. Other diet components such as wheat could be more important promoters of Type I diabetes.